RESUMO
BACKGROUND: Proximal tubule epithelial cells (PTECs) release proinflammatory and profibrogenic mediators when exposed to serum albumin that may contribute to progression of kidney disease. Interleukin 6 (IL-6) may influence renal fibrosis by modulating transforming growth factor beta1 (TGFbeta1) signalling. PTECs have been demonstrated to produce IL-6 in response to albumin treatment, but the mechanism has not been investigated. We hypothesized that albumin would induce release of IL-6 from PTECs, which would be sensitive to inhibition of PI3K, ERK1,2, p38 MAPK and NFkB. METHODS: Primary human PTECs were exposed to albumin (0.75-150 micronM) for 8 and 24 hours. IL-6 release was determined using enzyme-linked immunosorbent assay (ELISA). The effects of LY294002 (10 micronM), NH4Cl (10 mM), pyrrolidine dithiocarbamate (PDTC) (20 micronM), CAPE (17.5 micronM), PD098059 (20 micronM), SB202190 (5 micronM) and MG132 (10 micronM) on albumin-mediated IL-6 release were studied. RESULTS: Albumin caused a significant time- and concentration-dependent increase in IL-6 release by PTECs. LY294002, NH4Cl, CAPE, PD098059 and SB202190 all reduced albumin-mediated IL-6 release, but neither PDTC nor MG132 had any effect. CONCLUSIONS: These data demonstrate that albumin induces IL-6 release by primary human PTECs, and support a role for endocytosis, p38 MAPK, ERK1,2 and in this process.
Assuntos
Células Epiteliais/metabolismo , Interleucina-6/metabolismo , Túbulos Renais Proximais/metabolismo , Albumina Sérica/farmacologia , Antioxidantes/farmacologia , Proteínas Quinases Dependentes de Cálcio-Calmodulina/antagonistas & inibidores , Células Cultivadas , Cromonas/farmacologia , Inibidores Enzimáticos/farmacologia , Ensaio de Imunoadsorção Enzimática , Células Epiteliais/citologia , Células Epiteliais/efeitos dos fármacos , Flavonoides/farmacologia , Humanos , Imidazóis/farmacologia , Túbulos Renais Proximais/citologia , Túbulos Renais Proximais/efeitos dos fármacos , Proteína Quinase 1 Ativada por Mitógeno/antagonistas & inibidores , Morfolinas/farmacologia , Inibidores de Fosfoinositídeo-3 Quinase , Prolina/análogos & derivados , Proteínas Serina-Treonina Quinases/antagonistas & inibidores , Piridinas/farmacologia , Tiocarbamatos , Proteínas Quinases p38 Ativadas por Mitógeno/antagonistas & inibidores , Quinase Induzida por NF-kappaBRESUMO
Proteinuria predicts the decline of renal function in chronic kidney disease. Reducing albuminuria has been shown to be associated with a reduction in this rate of decline. Proximal tubular epithelial cells (PTECs), when exposed to albumin produce matrix proteins, proinflammatory and profibrotic cytokines like TGF-beta(1). Some of these effects are dependent on endocytosis of albumin by PTECs. However, conditions like diabetic nephropathy, believed to be associated with reduced albumin endocytosis, are associated with interstitial fibrosis. Moreover, megalin, the putative albumin binding receptor in PTECs, has potential signaling motifs in its cytoplasmic domain, suggesting its ability to signal in response to ligand binding from the apical surface of PTECs. Hence, we looked to see whether albumin-induced secretion of TGF-beta(1) by PTECs is dependent on albumin endocytosis or whether it could occur in the absence of albumin endocytosis. We studied the production of TGF-beta(1) in two accepted models of PTECs, opossum kidney cells and human kidney cell clone-8 cells, with widely varying degrees of endocytosis. We then studied the effect of inhibiting albumin endocytosis with various inhibitors on albumin-induced TGF-beta(1) secretion. Our results indicate that albumin-induced TGF-beta(1) secretion by PTECs does not require albumin endocytosis and therefore the mechanism for the induction of some profibrotic responses by albumin may differ from those required for some of the inflammatory responses. Moreover, we found that albumin-induced TGF-beta(1) secretion by PTECs is not dependent on its interaction with megalin.